WO2016160706A1 - Inhibiteurs de la collagène prolyl-4-hydroxylase - Google Patents

Inhibiteurs de la collagène prolyl-4-hydroxylase Download PDF

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WO2016160706A1
WO2016160706A1 PCT/US2016/024522 US2016024522W WO2016160706A1 WO 2016160706 A1 WO2016160706 A1 WO 2016160706A1 US 2016024522 W US2016024522 W US 2016024522W WO 2016160706 A1 WO2016160706 A1 WO 2016160706A1
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alkyl
groups
hydrogen
mmoles
carbon atoms
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Ronald Raines
James VASTA
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Wisconsin Alumni Research Foundation
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/78—Carbon atoms having three bonds to hetero atoms, with at the most one bond to halogen, e.g. ester or nitrile radicals
    • C07D213/79—Acids; Esters
    • C07D213/80—Acids; Esters in position 3
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
    • C07D413/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings directly linked by a ring-member-to-ring-member bond

Definitions

  • Collagen is the principle component of bone, connective tissues, and the extracellular matrix in animals. 1
  • the overproduction of collagen is associated with a variety of diseases, including fibrotic diseases 2 and cancers. 3"7
  • the stability of collagen relies on posttranslational modifications that occur throughout the secretory pathway. 8
  • the most prevalent of these modifications is the hydroxylation of collagen strands by collagen prolyl 4-hydroxylases (CP4Hs), which are Fe(II)- and oc-ketoglutarate (AKG)-dependent dioxygenases (FAKGDs) located in the lumen of the endoplasmic reticulum.
  • CP4Hs collagen prolyl 4-hydroxylases
  • AKG oc-ketoglutarate
  • FAKGDs oc-ketoglutarate
  • CP4Hs Catalysis by CP4Hs convert (25)-proline (Pro) residues in protocollagen strands into (25,4R)-4-hydroxyproline (Hyp) residues ( Figure 1A), which are essential for the conformational stability of mature collagen triple helices. 10 Importantly, CP4Hs are validated targets for treating both fibrotic diseases 11 and metastatic cancer, particularly breast cancer. 6
  • Fe(II) is bound by a conserved His-X- Asp/Glu...Xn...His motif, and AKG chelates to enzyme-bound Fe(II) using its C-l carboxylate and C-2 keto groups, while the C-5 carboxylate group forms hydrogen bonds and engages in Coulombic interactions with a basic residue (typically arginine or lysine).
  • CP4H inhibitors Like many FAKGDs, human CP4Hs are inhibited by simple metal chelators, such as 2,2'-bipyridine (bipy), as well as AKG mimics 18 , such as N-oxalyl glycine (NOG), pyridine-2,4-dicarboxylic acid (24PDC), and pyridine-2,5-dicarboxylic acid (25PDC), and 3,4-dihydroxybenzoic acid (DHB) 17 , as well as by simple metal chelators, such as 2,2'-bipyridine (bipy; Figure IB).
  • simple metal chelators such as 2,2'-bipyridine (bipy; Figure IB).
  • bipyridinedicarboxylic acids Two bipyDCs have high potency: 2,2'-bipyridine-4,5'-
  • bipy45'DC dicarboxylic acid
  • bipy55'DC 2,2'-bipyridine-5,5'-dicarboxylic acid
  • the bipyDCs represent an interesting class of compounds for the development of antifibrotic or antimetastatic therapeutics.
  • these compounds possess a variety of undesirable chemical properties that have limited their development thus far.
  • the bipyDCs as a class are not cell permeable, requiring the preparation of suitable cell permeable prodrugs.
  • the bipyDCs are capable of binding and forming complexes with free iron 38 and likely other biologically relevant metals.
  • inhibitors of CP4H and particularly those that have high potency and selectivity for CP4H compared to other P4Hs.
  • selective inhibitors of human CP4H There is also a need for such inhibitors which exhibit reduced iron binding while retaining inhibitor activity and/or which are cell permeable.
  • the invention provides biheteroaryl dicarboxylates and esters, and salts thereof which are useful generally as modulators of CP4H activity, more particularly as inhibitors of CP4H, or further as chemical genetic probes for the study of CP4H function and/or as synthetic intermediates for the preparation of such modulators, inhibitors and/or chemical genetic probes.
  • the invention provides compounds of formula I:
  • X is S, O, NH, or NR
  • R is an alkyl group having 1-3 carbon atoms
  • Ri and R2 independently are -OR7, or -NHSO2R8,
  • R 7 is selected from:
  • Rg is selected from hydrogen, alkyl, aryl, arylalkyl
  • R 3 , R4 and R 6 independently are hydrogen, alkyl, alkoxy, alkenyl, alkenoxy, haloalkyl, haloalkenyl, halogen, hydroxyl, hydroxyalkyl, hydroxyalkenyl, aryl, aryloxy, arylalkyl or arylalkyloxy;
  • R5 is hydrogen, halogen, alkyl having 1-3 carbon atoms, or alkoxy having 1-3 carbon atoms; -R'- is a divalent straight chain or branched alkylene, and
  • -R" is an alkyl, alkenyl, arylalkyl, or aryl group.
  • Alkyl, alkoxy, alkenyl, alkenyloxy, alkylene, arylalkyl, arylalkoxy, aryl or aryloxy groups are unsubstituted or are optionally substituted as defined herein below. In specific embodiments, these groups are substituted with one or more halo, hydroxyl, alkyl, alkoxy, haloalkyl, haloalkoxy, hydroxyalkyl, or hydroxyalkoxy groups.
  • Alkyl groups of substituent groups preferably have 1-3 carbon atoms.
  • Halogens include fluorine, chlorine, bromine and iodine. A specific halogen is fluorine. Another specific halogen is chlorine.
  • Haloalkyl and haloalkoxy groups can be monohalogenated through perhalogenated.
  • a specific haloalkyl group is trifluoromethyl.
  • a specific haloalkoxy group is trifluoromethoxy.
  • Hydroxyalkyl and hydroxyalkoxy groups can contain one or more hydroxyl groups, but more specifically contain 1, 2 or 3 hydroxyl groups.
  • X is S.
  • X is O.
  • X is NH
  • X is NCH 3 .
  • R 3 , R4, and R 6 are hydrogen, halogen or alkyl groups having 1-3 carbon atoms.
  • R 3 , R4, and R 6 are hydrogen, halogen, phenyl, benzyl or phenethyl groups, where phenyl, benzyl and phenethyl groups are unsubstituted or substituted with one or more non-hydrogen substituents.
  • Specific substituents for phenyl, benzyl and phenethyl groups are halogens, hydroxyl groups, alkyl groups having 1-3 carbon atoms or alkoxy groups having 1-3 carbon atoms.
  • R 3 , R4, and R 6 are hydrogen or alkyl groups having 1-3 carbon atoms.
  • R 3 , R4, and R 6 are hydrogens.
  • R5 is hydrogen, methyl or methoxy.
  • R5 is methyl or methoxy.
  • R5 is methyl
  • R5 is hydrogen
  • R 3 , R4, and R 6 are hydrogen, halogen, alkyl having 1-3 carbon atoms or alkoxy having 1 -3 carbon atoms and R5 is hydrogen.
  • Ri and R2 are independently -OR7 groups where R7 is hydrogen or an alkyl group having 1-8 carbon atoms.
  • Ri and R2 are independently -OR7 groups where R7 is an alkyl group having 1-8 carbon atoms.
  • Ri and R2 are independently -OR7 groups where R7 is an alkyl group having 1-3 carbon atoms.
  • Ri and R2 are -OR7 groups where R7 is a methyl group.
  • Ri and R2 are methoxy groups
  • R5 is hydrogen and R 3 , R4 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy.
  • Ri and R2 are methoxy groups
  • R 3 , R4, R5 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy.
  • Ri and R2 are ethyl groups. In specific embodiments of formula I, Ri and R2 are ethoxy groups, R5 is hydrogen and R 3 , R4 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy. In specific embodiments of formula I, Ri and R2 are ethoxy groups, and R 3 , R4, R5 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy. In specific embodiments of formula I, Ri and R2 are -0-(CH2) n -0-CO-R" groups, where n is 1-3 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R2 are -O- (CH2)n-0-CO-R" groups, where n is 1-3 and R" is an alkyl having 1-4 carbon atoms. In specific embodiments, Ri and R2 are -0-(CH2) n -0-CO-R" groups, where n is lor 2 and R" is an alkyl having 1-6 carbon atoms. In specific embodiments, Ri and R2 are -0-(CH2)-0-CO- R' groups where R" is an alkyl having 1 -4 carbon atoms.
  • Ri and R2 are -0-(CH2)-0-CO-R" groups, where R' is a methyl, ethyl, n-propyl or iso-propyl, n-butyl, iso-butyl, sec-butyl or t-butyl group.
  • Ri and R2 are -(CH2)-0-CO- R" groups, where R" is t-butyl.
  • Ri and R2 are -0-(CH2) n -0-CO-0-R" groups, where n is 1 -3 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R2 are - 0-(CH2) n -0-CO-0-R" groups, where n is 1-3 and R" is an alkyl having 1-4 carbon atoms.
  • Ri and R2 are -0-(CH2) n -0-CO-0-R" groups, where n is lor 2 and R" is an alkyl having 1 -6 carbon atoms.
  • Ri and R2 are -0-( ⁇ 3 ⁇ 4)-0- CO-O-R" groups, where R" is an alkyl having 1-4 carbon atoms.
  • Ri and R2 are -0-(CH2) n -0-CO-0-R" groups, where R" is a methyl, ethyl, n-propyl or iso- propyl, n-butyl, iso-butyl, sec-butyl or t-butyl group.
  • Ri and R2 are -0-(CH 2 )-0-CO-0-R" groups, where R" is iso-propyl.
  • one of Ri or R2 is a -NHSO2R8 group.
  • Ri is a -NHSO2R8 group and R2 is a -OR7 group.
  • R2 is a -NHSO2R 8 group and Ri is a -OR7 group.
  • R7 is hydrogen.
  • R7 is an alkyl group having 1-8 carbon atoms.
  • Rs is hydrogen. In specific embodiments, Rs is an alkyl group having 1 -6 carbon atoms. In specific embodiments, Rs is an alkyl group having 1-3 carbon atoms. In specific embodiments, Rs is an unsubstituted phenyl group. In specific embodiments, Rs is an unsubstituted benzyl group. In specific embodiments, Rs is an unsubstituted phenethyl group. In specific embodiments, Rs is a substituted phenyl, benzyl or phenethyl group, having 1 , 2 or 3 non-hydrogen substituents, which include among others halogen, hydro yl, alkyl or alkoxy groups.
  • Ri and R2 are -OR7 groups where R7 is an alkyl or an alkoxyalkyl.
  • R7 is an unsubstituted alkyl or an alkyl group substituted with one or more halogens.
  • R7 is an alkoxyalkyl group having a total of 2-8 carbon atoms. One or more carbons of the alkoxyalkyl group can be substituted with one or more halogens.
  • the invention provides methods for inhibition of CP4H by contacting the CP4H with a compound of formula I or a salt thereof.
  • the method employs a compound of formula I where X is S.
  • the method employs a compound of formula I where X is O.
  • the method employs a compound of formula I where X is NH.
  • the method employs a compound of formula I where X is NCH 3 .
  • Ri and R2 are -OR7, where R7 is an alkyl group having 1-6 carbon atoms, or a alkyl group having 2-6 carbon atoms, or a alkyl group having 1-3 carbon atoms, or a alkyl group having 1-3 carbon atoms.
  • the invention is also directed to pharmaceutical compositions comprising a therapeutically effective amount of one or more of the compounds of formula I or salts thereof and a pharmaceutically acceptable carrier.
  • Such pharmaceutical compositions are useful for inhibition of CP4H in an individual in need of such inhibition.
  • Compounds of formula I and pharmaceutical compositions comprising them are useful as antifibrotic agents.
  • Compounds of formula I and pharmaceutical compositions comprising them are useful for treatment of fibrotic diseases and disorders.
  • Compounds of formula I and pharmaceutical compositions comprising them are useful for treatment of fibrotic liver disease, idiopathic fibrosis, pulmonary fibrosis, renal fibrosis, cardiac fibrosis, and fibrosis associated with scleroderma or rheumatoid arthritis.
  • Compounds of formula I and pharmaceutical compositions comprising them are useful for treatment of diseases and disorders involving undesirable collage deposition.
  • the invention is also directed to cellular inhibitors of CP4H which are useful as research reagents in experiments in cultured cells which can be used for inhibition at concentrations that do not cause iron deficiency.
  • Figure 1 A illustrates the reaction catalyzed by collagen prolyl 4-hydroxylase (CP4H) and its inhibition.
  • CP4Hs catalyze the hydroxylation of Pro residues in collagenous peptides to form Hyp residues.
  • Figure IB shows chemical structures of examples of previously reported human CP4H inhibitors.
  • Figure 2 shows structures of biheteroaryl compounds used in this work.
  • Figure 3 A is a graph of ligand titrations to form iron complexes for biheteroaryl compounds as indicated. Absorbance at ax is measured as a function of compound concentration. R 2 >
  • Figure 3B is a graph of ligand titrations to form iron complexes for diethyl pyimDC.
  • Fe(II) (20 ⁇ ) could not be saturated over the range that diethyl pyimDC remained soluble under the assay conditions. Thus, Fe2o-EC5o >600 ⁇ , which was the highest concentration tested.
  • Figure 4 illustrates the results of a screen for inhibition of human CP4H1. Compounds (10 ⁇ ) were screened for inhibition of the catalytic activity of human CP4H1 as described in The Examples. Relative activity values are the mean ( ⁇ SD) of three replicates. Data for 25PDC, bipy45'DC, and bipy55'DC are from Vasta et al, 2015 38 .
  • Figure 5A is a graph illustrating competitive iron-binding by pyimDC with bipy. High concentrations of pyimDC can compete with bipy for complexation with Fe(II), as determined with the assay described in the Examples. These data are consistent with pyimDC forming the depicted 2:1 complex with iron. This phenomenon was dose-dependent and confined to the diacids (not the corresponding esters).
  • Figure 5B is a table illustrating structures of NMe-PyinDC, PyimDC and its corresponding methyl and ethyl diesters and providing EC5 0 for complex if observed.
  • Figure 6A is a graph showing biheteroaryl dicarboxylates as inhibitors of human CP4H1. Individual points represent the mean ( ⁇ SD) of three independent experiments. Data were fitted to a dose-response equation to determine IC50 values: pyimDC, (2.6 + 0.1) ⁇ ;
  • Figure 6B illustrates Lineweaver-Burke analysis of inhibition by pythiDC.
  • the rate of the reaction catalyzed by CP4Hlwith increasing a-ketoglutarate concentration (10-100 ⁇ ) was determined in the presence of a fixed concentration of pythiDC (0.0,
  • Figure 7 illustrates the results of a screen for inhibition of human PHD2.
  • Compounds (10 ⁇ ) were screened for inhibition of the catalytic activity of human PHD2 as described in The Examples. Relative activity values are the mean ( ⁇ SD) of three replicates. Data for NOG, 24PDC, 25PDC, bipy, bipy45'DC, and bipy55'DC are from Vasta et al. 38
  • Figure 8A illustrates the effect of esterified biheteroaryl compounds on iron metabolism. MDA-MB-231 breast cancer cells were treated with deferoxamine (DFO), ethyl
  • DFO deferoxamine
  • Figure 8B are densitometric quantitations (n > 3) corresponding to the blots in Figure 8A and normalized to ⁇ -actin.
  • the dose of EDHB (500 ⁇ ) is known to diminish collagen secretion significantly.
  • 6 Relative ferritin levels are shown in the left graph.
  • Relative TfR levels are shown in the right graph.
  • Figure 8C illustrates the effect of esterified biheteroaryl compounds on collagen secretion into conditioned media. Blots (top) are representative of at least five replicates and densitomeric quantitations (bottom) are normalized to total protein using the Ponceau S- stained blot.
  • Figure 9A illustrates the effect of biheteroaryl compounds on iron metabolism in human breast cancer cells, (a) MDA-MB-231 breast cancer cells were treated with deferoxamine (DFO), biheteroaryl compounds, or vehicle (DMSO) as described in the Examples, and analyzed with an immunoblot (*, p ⁇ 0.05).
  • DFO deferoxamine
  • DMSO vehicle
  • Figure 9B are densitometric quantitations corresponding to the blots in Figure 9A.
  • Relative ferritin levels are shown in the left graph.
  • Relative TfR levels are shown in the right graph.
  • Figure 10 illustrates the effect of biheteroaryl compounds and their diethyl esters on iron metabolism in human embryonic kidney cells.
  • HEK293T cells were treated with biheteroaryl compounds (100 ⁇ ), deferoxamine (DFO, 100 ⁇ ), or vehicle (DMSO) as described in the Examples, and analyzed with an immunoblot.
  • DFO deferoxamine
  • DMSO vehicle
  • FIG 11 illustrates the effect of esterified biheteroaryl compounds on IRE Binding by IRPs in Human Breast Cancer Cells.
  • MDA-MB-231 cells were treated with esterified biheteroaryl compounds, deferoxamine (DFO), ethyl dihydroxybenzoate (EDHB), or vehicle (DMSO) as described in the Examples, and analyzed by an electrophoretic mobility shift assay (EMSA) using a 32 P- labeled RNA ligand for IRPs.
  • DFO deferoxamine
  • EDHB ethyl dihydroxybenzoate
  • DMSO vehicle
  • MDA-MB-231 cells were treated with esterified biheteroaryl compounds, deferoxamine (DFO), ethyl dihydroxybenzoate (EDHB), or vehicle (DMSO) as described in the Examples, and then analyzed with an immunoblot. Blots are representative of at least 2 replicates.
  • DFO deferoxamine
  • EDHB ethyl dihydroxybenzoate
  • DMSO vehicle
  • the invention is based at least in part on the preparation and investigation of certain bihe X:
  • R -R are as defined for formula I and HET is selected from:
  • R5 is hydrogen, halogen, alkyl having 1-3 carbon atoms, or alkoxy having 1-3 carbon atoms and R 6 is as defined for formula I;
  • R 6 is as defined for formula I and R is hydrogen or an alkyl having 1 -3 carbons atoms and is specifically hydrogen or is specifically methyl; where R is hydrogen or an alkyl group. R is more specifically hydrogen or methyl.
  • Certain compounds of formula X modulate the activity of collagen prolyl 4-hydroxylases. Certain compounds of formula X modulate the activity of human collagen prolyl 4- hydroxylase. Certain compounds of formula X inhibit the activity of collagen prolyl 4- hydroxylases. Certain compounds of formula X inhibit the activity of human collagen prolyl 4-hydroxylase. Certain compounds of formula X increase the activity of collagen prolyl 4- hydroxylases. Certain compounds of formula X increase the activity of human collagen prolyl 4-hydroxylase.
  • HET is Al.
  • HET is A2.
  • HET is Bl.
  • HET is B2.
  • HET is B3.
  • HET is CI
  • HET is C2.
  • X is NR, where R is an alkyl group having 1-3 carbon atoms or more specifically R is methyl.
  • Y is C and Z is N.
  • R is methyl.
  • R 3 , R4, and R 6 are hydrogen, halogen or alkyl groups having 1-3 carbon atoms.
  • R 3 , R4, and R 6 are hydrogen, halogen, phenyl or benzyl groups, where phenyl and benzyl groups are
  • R 3 , R4, and R 6 are hydrogen or alkyl groups having 1-3 carbon atoms.
  • R 3 , R4, and R 6 are hydrogens.
  • R5 is hydrogen, methyl or methoxy.
  • R5 is methyl or methoxy.
  • R5 is methyl
  • R5 is hydrogen.
  • R 3 , R4, and R 6 are hydrogen, halogen, alkyl having 1-3 carbon atoms or alkoxy having 1-3 cabon atoms and R5 is hydrogen.
  • Ri and R2 are independently -OR7 groups where R7 is hydrogen or an alkyl group having 1-8 carbon atoms. In specific embodiments of formula X, and for each embodiment of HET, Ri and R2 are independently -OR7 groups where R7 is an alkyl group having 1-8 carbon atoms.
  • Ri and R2 are independently -OR7 groups where R7 is an alkyl group having 1-3 carbon atoms.
  • Ri and R2 are -OR7 groups where R7 is a methyl group.
  • R5 is hydrogen and R 3 , R4 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy.
  • Ri and R2 are methoxy groups, and R 3 , R4, R5 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy.
  • Ri and R2 are ethyl groups. In specific embodiments of formula X, and for each embodiment of HET, Ri and R2 are ethoxy groups, R5 is hydrogen and R 3 , R4 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy. In specific embodiments of formula X, and for each embodiment of HET, Ri and R2 are ethoxy groups, and R 3 , R4, R5 and R 6 are hydrogen, halogen, hydroxyl, methyl, or methoxy.
  • Ri and R2 are -O- (CH2)n-0-CO-R" groups where n is 1-3 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )n-0-CO-R" groups where n is 1-3 and R" is an alkyl having 1-4 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )n-0-CO-R" groups where n is lor 2 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )-0-CO-R' groups where R" is an alkyl having 1-4 carbon atoms. In specific embodiments, Ri and R 2 are -0-(CH 2 )-0-CO-R" groups where R' is a methyl, ethyl, n-propyl or iso-propyl, n-butyl, iso-butyl, sec-butyl or t-butyl group. In specific embodiments, Ri and R 2 are -(Cty-O-CO-R" groups where R" is t-butyl.
  • Ri and R 2 are -O- (CH 2 )n-0-CO-0-R" groups where n is 1-3 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )n-0-CO-0-R" groups where n is 1-3 and R" is an alkyl having 1-4 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )n-0-CO- O-R" groups where n is lor 2 and R" is an alkyl having 1-6 carbon atoms.
  • Ri and R 2 are -0-(CH 2 )-0-CO-0-R" groups where R" is an alkyl having 1-4 carbon atoms. In specific embodiments, Ri and R 2 are -0-(CH 2 )n-0-CO-0-R" groups where R" is a methyl, ethyl, n-propyl or iso-propyl, n-butyl, iso-butyl, sec-butyl or t-butyl group. In specific embodiments, Ri and R 2 are -0-(CH 2 )-0-CO-0-R" groups where R' is iso-propyl.
  • Ri or R 2 is a -NHSO 2 R 8 group.
  • Ri is a -NHSO 2 R 8 and R 2 is a -OR 7 group.
  • R 2 is a -NHSO 2 R 8 and Ri is a hydrogen or a -OR 7 group.
  • R 7 is hydrogen.
  • R 7 is an alkyl group having 1-8 carbon atoms.
  • Rs is hydrogen.
  • Rs is an alkyl group having 1-6 carbon atoms.
  • Rs is an alkyl group having 1- 3 carbon atoms.
  • Rg is an unsubstituted phenyl group. In specific embodiments, Rg is an unsubstituted benzyl group. In specific embodiments, Rg is an unsubstituted phenethyl group.
  • Ri and R 2 are - OR 7 groups where R 7 is alkyl or alkoxyalkyl.
  • the invention provides methods for modulation of CP4H by contacting the CP4H with a compound of formula X or a salt thereof.
  • the method employs a compound of formula X where Het is Al , Bl, B2, B3, CI or C2.
  • alkyl or alkyl group refer to a monoradical of a straight-chain or branched saturated hydrocarbon.
  • Alkyl groups include straight-chain and branched alkyl groups. Unless otherwise indicated alkyl groups have 1-12 carbon atoms (C1-C12 alkyl groups) and preferred are those that contain 1-8 carbon atoms (C1-C8 alkyl groups), more preferred are those that contain 1-6 carbon atoms (C1-C6 alkyl groups) and yet more preferred are those that contain 1-3 carbon atoms (C1-C3 alkyl groups).
  • Alkyl groups are optionally substituted with one or more non-hydrogen substituents as described herein.
  • alkyl groups include haloalkyl groups and hydroxyalkyl groups.
  • exemplary alkyl groups include methyl, ethyl, n-propyl, iso-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, branched-pentyl, n- hexyl, branched hexyl, all of which are optionally substituted.
  • Substituted alkyl groups include fully halogenated or semihalogenated alkyl groups, such as alkyl groups having one or more hydrogens replaced with one or more fluorine atoms, chlorine atoms, bromine atoms and/or iodine atoms.
  • Substituted alkyl groups include fully fluorinated or semifluorinated alky groups.
  • Substituted alkyl groups include alkyl groups substituted with one or more hydroxyl groups. Particular hydroxyalkyl groups are those having 1, 2, 3 or 4 hydro xyl groups and particularly those having one hydroxyl group.
  • the term alkyl group further includes cycloalkyl groups which are those alkyl groups which have 1 ring or which are bicyclic or tricyclic. In specific embodiments, cycloalkyl groups have one ring having 5-8 carbon atoms and preferably have one 5-or 6-carbon ring. Alkyl groups can be unsubstituted. Alkyl groups can be other than cycloalkyl groups.
  • alkoxy group is an alkyl group, as broadly discussed above, linked to oxygen (-O-R a i k yi)-
  • alkoxyalkyl refers to a group having two or more alkylene or alkyl moieites each linked through oxygen atoms, e.g., -alkylene-0-alkyl, -alkylene-O-alkylene-O-alkyl, etc.or more generically -0-(alkylene-0)n-alkyl, where n is an integer from 1 to 20.
  • Alkylene groups can contain 1-12 carbon atoms or more specifically 1-6 carbon atoms or more specifically 1-3 carbon atoms.
  • Alkyl groups can contain 1-12 carbon atoms, 1-6 carbon atoms or 1-3 carbon atoms. Alkoxyalkyl groups can contain 1-22 carbon atoms and 1-10, 1-6 or 1-3 oxygen atoms. Such groups can also be designated ether groups.
  • the alkylene moiety can be straight-chain or branched. The alkyl group can be straight-chain or branched.
  • alkoxyalkyl groups include: -(CH2-0) n -CH3, where n is 1-6 or 1-3, -(CH 2 CH 2 -0) n - CH 3 , where n is 1-6 or 1-3, or -(CH 2 CH 2 CH2-0)n-CH3, where n is 1-6 or 1-3, -(CH 2 )n-0- (CH 2 -CH(Ra)-CH 2 )-0-(CH 2 ) m -CH 3 , where n is 1-6, m is 1-6 and Ra is an alkyl having 1-4 carbon atoms.
  • alkenyl or alkenyl group refer to a monoradical of a straight-chain or branched hydrocarbon having one or more double bonds. Unless otherwise indicated alkenyl groups have 2-12 carbon atoms (C2-C12 alkenyl groups) and preferred are those that contain 2-8 carbon atoms (C1-C8 alkenyl groups), more preferred are those that contain 2-6 carbon atoms (C2-C6 alkyl groups) and yet more preferred are those that contain 2-3 carbon atoms (C2-C3 alkyl groups).
  • Alkenyl groups are optionally substituted with one or more non-hydrogen substituents as described herein. Specific substituted alkenyl groups include haloalkenyl groups and hydroxyalkenyl groups.
  • alkenyl groups include ethenyl, n-propenyl, iso-propenyl, n-butenyl, s-butenyl,n-pentenyl, branched-pentenyl, n-hexenyl, branched hexenyl, all of which are optionally substituted.
  • Substituted alkenyl groups include fully halogenated or semihalogenated alkenyl groups, such as alkenyl groups having one or more hydrogens replaced with one or more fluorine atoms, chlorine atoms, bromine atoms and/or iodine atoms.
  • Substituted alkenyl groups include fully fluorinated or semifluorinated alkenyl groups.
  • Substituted alkenyl groups include alkenyl groups substituted with one or more hydroxyl groups. Particular hydroxyalkenyl groups are those having 1, 2, 3 or 4 hydroxyl groups and particularly those having one hydroxyl group.
  • the term alkenyl group further includes cycloalkenyl groups which are those alkenyl groups which have 1 ring or which are bicyclic or tricyclic. In specific embodiments, cycloalkenyl groups have one ring having 5-8 carbon atoms and preferably have one 5 -or 6-carbon ring. Specific alkenyl groups are those having one double bond. Specific alkenyl groups are those having 2 double bonds.
  • Alkenyloxy refers to an alkenyl group linked through oxygen (-O-Rai k enyi)-
  • Alkylene refers to divalent moieties derived formally from alkyl groups as described above by removal of an additional hydrogen e.g., -(C]3 ⁇ 4) a -, where a is 1-20.
  • Alkylene moieties are optionally branched or substituted, e.g., -(CH 2 ) a -CH(CH 3 )-(CH 2 )t,-, where the sum of a + b is 1 to 20.
  • Alkylene groups can have 1-20, 1-10, 1-6 or 1-3 carbon atoms.
  • Alkylene groups can have 1, 2, 3, 4, 5 or 6 carbon atoms.
  • Aryl groups include groups having one or more 5- or 6-member aromatic rings.
  • Aryl groups can contain one, two or three, 6-member aromatic rings.
  • Aryl groups can contain two or more fused aromatic rings.
  • Aryl groups can contain two or three fused aromatic rings.
  • Aryl groups are optionally substituted with one or more non-hydrogen substituents.
  • Substituted aryl groups include among others those which are substituted with alkyl or alkenyl groups, which groups in turn can be optionally substituted.
  • Specific aryl groups include phenyl groups, biphenyl groups, pyridinyl groups, and naphthyl groups, all of which are optionally substituted as described herein.
  • Aryl groups include heteroaryl groups having 1-3 heteroatoms in the one or more 5- or 6-member rings.
  • Heteroatoms include O, S and N.
  • Aryl groups include those having only carbons in the rings.
  • Substituted aryl groups include fully halogenated or semihalogenated aryl groups, such as aryl groups having one or more hydrogens replaced with one or more fluorine atoms, chlorine atoms, bromine atoms and/or iodine atoms.
  • Substituted aryl groups include fully fluorinated or semifluorinated aryl groups, such as aryl groups having one or more hydrogens replaced with one or more fluorine atoms.
  • Specific aryl groups are unsubstituted and substituted phenyl groups.
  • Specific aryl rings include unsubstituted or substituted naphthyl groups.
  • Specific aryl groups are halogenated phenyl groups.
  • An aryloxy group is an aryl group, as broadly discussed above, linked to oxygen (-O-Raryi).
  • Arylalkyl groups are alkyl groups substituted with an aryl group. Specific arylalkyl groups are benzyl (-CH2-phenyl) or phenethyl (-CH 2 -CH 2 -phenyl).
  • Arylalkoxy groups are arylalkyl groups linked to oxygen (-O-Raryiaikyi)
  • Groups e.g., alkyl, alkylene, aryl, arylalkyl, alkenyl, phenyl and benzyl
  • groups herein are optionally substituted most generally, for example, with one or more oxo group, thioxo group, halogen, nitro, cyano, cyanate, azido, thiocyano, isocyano, isothiocyano, sulfhydryl, hydroxyl, alkyl, alkoxy, alkylthio, -COR s , -COH, -OCOR s , -OCOH, -CO-OR s , -CO-OH, -CO-0-CO-R s , -
  • optional substitution is substitution with 1-12 non-hydrogen substituents. In specific embodiments, optional substitution is substitution with 1-6 non- hydrogen substituents. In specific embodiments, optional substitution is substitution with 1-3 non-hydrogen substituents. In specific embodiments, optional substituents contain 6 or fewer carbon atoms. In specific embodiments, optional substitution is substitution by one or more halogen, hydroxy group, cyano group, oxo group, thioxo group, unsubstituted C1-C6 alkyl group or an unsubstituted aryl group.
  • any of the above groups which contain one or more substituents it is understood, that such groups do not contain any substitution or substitution patterns which are sterically impractical and/or synthetically non-feasible.
  • the compounds of this invention include all stereochemical isomers arising from the substitution of these compounds.
  • the invention provides methods for modulating the activity of collagen prolyl 4-hydoxylase activity in vitro and in vivo. Modulation of collagen prolyl 4-hydoxylase activity facilitates modulation of the biosysnthesis of collagen.
  • the invention provides a method for modulating the biosynthesis of collagen.
  • CP4H activity is modulated by contact thereof with one or more compounds of formula X.
  • in vivo inhibition is inhibition in cells.
  • the invention provides methods for inhibiting the activity of collagen prolyl 4-hydoxylase activity in vitro and in vivo. Inhibition of collagen prolyl 4-hydoxylase activity facilitates modulation of the biosysnthesis of collagen.
  • the invention provides a method for inhibiting the biosynthesis of collagen. CP4H activity is inhibited by contact thereof with one or more compounds of formula X.
  • Fibrosis can affect any organ or tissue in the body and include fibroses of the lung, liver, skin and atherosclerosis. Fibrotic diseases and disorders also include scleroderma and scarring after burns, or other injuries or after surgery. Fibrotic diseases also include fibroproliferative disorders, including liver cirrhosis, pulmonary fibrosis, and systemic sclerosis, among others. Collagen is deposited during fibrosis. Inhibition of collagen prolyl 4-hydroxylase results in inhibition of collagen biosynthesis, which reduces undesired collagen deposition.
  • Gilkes et al. 6 reports that the presence of hypoxia and fibrosis within the primary tumor are two major risk factors for metastasis of human breast cancer. They report further that hypoxia-inducible factor 1 activates the transcription of genes encoding collagen prolyl hydroxylases. These genes function for collagen deposition by breast cancer cells. They report that expression of collagen prolyl hydroxylases promotes cancer cell alignment along collagen fibers, resulting in enhanced invasion and metastasis to lymph nodes and lungs. They report that ethyl 3 ,4-dihydroxybenzoate, a prolyl hydroxylase inhibitor, decreases tumor fibrosis and metastasis in a mouse model of breast cancer.
  • the present invention provides inhibitors of CP4H, for use in treatment of cancer, in particular breast cancer, for preventing or inhibiting metastasis of breast cancer. More specifically, the invention provides compounds of formula I for such treatment. Pharmaceutical compositions comprising one or more compounds of formula I in an amount effective for inhibition of CP4H in vivo for such treatment are provided. The compounds, compositions and methods of this invention can be combined if desired with art-known methods of treatment of cancer, particularly breast cancer.
  • the invention provides pharmaceutical compositions comprising a pharmaceutically effective amount of one or more compounds and /or salts of formula I and a pharmaceutically acceptable carrier or excipient.
  • the compounds and salts thereof of the invention can be used to prepare medicaments for the treatment and prevention of fibrotic diseases and disorders and the symptoms associated therewith.
  • pharmaceutically effective amount refers to an amount effective for treatment of a fibrotic disease or disorder in an individual (human or other mammal) in need of such treatment either by administration of a single compound or salt of formula I or in
  • the pharmaceutically effective amount of a given compound when administered as the only active ingredient may differ from its pharmaceutically effective amount when administered with other active ingredients. It will be appreciated that the pharmaceutically effective amount of a compound may differ from that of a salt of the same compound. Treating includes the alleviation of symptoms of a particular disorder in a patient or a measurable improvement of a parameter associated with a particular disorder. Treating includes treatment to prevent, delay or decrease undesired deposition of collagen.
  • prolactic ally effective amount refers to an amount of a compound or salt of the invention effective in preventing such deposition in an individual.
  • CP4H inhibitory effective amount' is that amount of a compound effective for inhibiting a given collagen prolyl 4-hydroxylase. The amount effective in vivo in an organism or in a cell may be different.
  • collagen inhibitory effective amount is that amount of a compound that is effective for inhibiting collagen biosynthesis.
  • the compounds and salts of formula I of the invention can be employed for treatment as is known in the art for other collagen prolyl 4-hydoxylase inhibitors as described in one or more of the patent documents cited herein and as is known in the art.
  • the term "individual” includes reference to a mammal, including a human.
  • Compounds of the invention of formula I can be administered in the form of
  • salts which include the following no n- limiting examples: alkali metal salts, such as those of lithium, potassium and sodium; alkali earth metal salts, such as those of barium, calcium and magnesium; transition metal salts, such as those of zinc; and other metal salts, such as those of aluminum, sodium hydrogen phosphate and disodium phosphate; salts of nitrates, borates, methanesulfonates, benzene sulfonates,
  • toluenesulfonates salts of mineral acids, such as those of hydrochlorides, hydrobromides, hydroiodides and sulfates; salts of organic acids, such as those of acetates, trifuoroacetates, maleates, oxalates, lactates, malates, tartrates, citrates, benzoates, salicylates, ascorbates, succinates, butyrates, valerates and fumarates.
  • amine salts such as those of ⁇ , ⁇ '- dibenzylethylenediamine, chloroprocaine, choline, ammonia, diethanolamine and other hydroxyalkylamines, ethylenediamine, N-methylglucamine, procaine, N- benzylphenethylamine, l-para-chlorobenzyl-2-pyrrolidin- 1 ' -ylmethyl-benzimidazole, diethylamine and other alkylamines, piperazine and tris(hydroxymethyl)- aminomethane.
  • Pharmaceutically acceptable salts can be derived from inorganic or organic acids or can be derived from inorganic or organic bases as is known in the art.
  • Basic amino acids useful for salt formation include arginine, lysine and ornithine.
  • Acidic amino acids useful for salt formation include aspartic acid and glutamic acid.
  • Compound of the invention can be administered in the form of pharmaceutically acceptable esters which include, among others, alkyl, alkenyl, alkynyl, aryl, heteroaryl, cycloalkyl and heterocyclyl esters of acidic groups, including, but not limited to, carboxylic acids, phosphoric acids, phosphinic acids, sulfonic acids, sulfinic acids and boronic acids.
  • compositions or dosage forms can be administered by any known route that is appropriate for the individual being treated and for the treatment or prophylaxis that is desired.
  • Specifically administration can be orally or non-orally in the form of, for example, granules, powders, tablets, capsules, syrup, suppositories, injections, emulsions, elixir, suspensions or solutions, by mixing these effective components, individually or simultaneously, with pharmaceutically acceptable carriers, excipients, binders, diluents or the like.
  • a solid formulation for oral administration can comprise one or more of the compounds or salts of the invention alone or in appropriate combination with other active ingredients.
  • Solid formulations can be in the form of powders, granules, tablets, pills and capsules.
  • the instant compounds can be mixed with at least one additive, for example, sucrose, lactose, cellulose sugar, mannitol, maltitol, dextran, starch, agar, alginates, chitins, chitosans, pectins, tragacanth gum, gum arabic, gelatins, collagens, casein, albumin, synthetic or semisynthetic polymers or glycerides.
  • formulations can contain, as in conventional cases, further additives, for example, an inactive diluent, a lubricant such as magnesium stearate, a preservative such as paraben or sorbic acid, an anti-oxidant such as ascorbic acid, tocopherol or cysteine, a disintegrator, a binder, a thickening agent, a buffer, a sweetener, flavoring agent and/or a perfuming agent.
  • Tablets and pills can also be prepared with enteric coating.
  • Standard methods of formulation can be applied to preparation of formulations of the compounds and salts of the invention.
  • Non-oral administration includes subcutaneous injection, intravenous injection, intramuscular injections, intraperitoneal injection or instillation.
  • injectable preparations for example, sterile injectable aqueous suspensions or oil suspensions can be prepared by known methods.
  • compositions may be formulated as known in the art for nasal aerosol or inhalation and may be prepared as solutions in saline, and benzyl alcohol or other suitable preservatives, absorption promoters, fluorocarbons, or solubilizing or dispersing agents.
  • Rectal suppositories can be prepared by mixing the drug with a suitable vehicle, for example, cocoa butter and polyethylene glycol, which is in the solid state at ordinary temperatures, in the liquid state at temperatures in intestinal tubes and melts to release the drug.
  • suitable vehicle for example, cocoa butter and polyethylene glycol
  • liquid preparations for oral administration include pharmaceutically acceptable emulsions, syrups, elixirs, suspensions and solutions, which may contain an inactive diluent, for example, pharmaceutically acceptable water.
  • the pharmaceutical composition can be formulated for topical administration, for example, with a suitable ointment containing one or more of the compounds or salts of the invention suspended or dissolved in a carrier, which include mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax and pharmaceutically acceptable water.
  • a suitable ointment containing one or more of the compounds or salts of the invention suspended or dissolved in a carrier, which include mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax and pharmaceutically acceptable water.
  • topical formulations can be formulated with a lotion or cream containing the active compound suspended or dissolved in a carrier.
  • Suitable carriers include mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and pharmaceutically acceptable water.
  • dosages of the instant compounds are dependent on age, body weight, general health conditions, sex, diet, dose interval, administration routes, excretion rate, combinations of drugs and conditions of the diseases treated. While taking these and other necessary factors into consideration, generally, dosage levels of between about 10 pg per day to about 5000 mg per day, preferably between about 100 mg per day to about 1000 mg per day of the compound are useful in the prevention and treatment of fibrotic diseases or disorders.
  • the pharmaceutical compositions of this invention will be administered from about 1 to about 5 times per day or alternatively, as a continuous infusion. Such administrationcan be used as a chronic or acute therapy.
  • the amount of active ingredient that may be combined with the carrier or excipient materials to produce a single dosage form will vary depending upon the patient/individual treated and the particular mode of administration.
  • a typical preparation will contain from about 5% to about 95% active compound (WAV).
  • WAV active compound
  • a typical preparation will contain from about 0.05% to about 95% active compound (WAV). Preferably, such preparations contain from about 10% to about 80% active compound.
  • WAV active compound
  • the desired unit dose of the composition of this invention is administered once or multiple times daily.
  • compounds of the invention include pharmaceutically acceptable salts or solvates thereof of formula X and particularly of formula I, which retain the physiologic activity of the corresponding free base or acid.
  • the salts and free base or acid forms of the compounds of the invention may be different in some physical properties, such as, solubility. See: for example, S. M. Berge, et al., "Pharmaceutical Salts," J. Pharm. Sci., 66: 1-19 (1977), which is incorporated by reference herein for teachings with respect to salts and solvates thereof.
  • the compounds of the present invention or pharmaceutically acceptable salts thereof can form solvates, such as hydrates, or alcoholates. Methods are known in the art for making solvates and particularly hydrates of compounds and salts. Salts of the invention can be in the form of solvates and particularly in the form of hydrates.
  • the compounds of any one of Formula I and salts and solvates thereof can be used in manufacture of a medicament for the treatment of fibrotic disorders or disorders including any disorder that is associated with undesirable collage deposition.
  • the invention provides use of one or more of the compounds of formula I as an antifibrotic agent.
  • solvate as used herein is a combination, or physical association of a compound with a solvent molecule.
  • a specific solvate is a hydrate.
  • Solvates can include those where the molar ratio of solvent to compound ranges, for example from 1 ⁇ 2 to 10 or more typically 1 ⁇ 2 to 4 and can include a disolvate, monosolvate or hemisolvate, among others. This physical association can involve varying degrees of ionic and covalent bonding, including hydrogen bonding.
  • the solvate can be isolated, such as when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid.
  • solvent encompasses both solution-phase and isolatable solvates.
  • solvates are isolatable with one or more molecules of solvent incorporated into the crystal lattice of a crystalline solid.
  • Compounds of the invention may be present as solvated forms with a pharmaceutically acceptable solvent, such as water, methanol, ethanol, and the like, and it is intended that the invention includes both solvated and unsolvated forms of compounds of the invention.
  • Solvates typically can function as pharmacological equivalents. Solvates typically do not significantly alter the physiological activity or toxicity of the compounds. Preparation of solvates is known in the art. See, for example, M. Caira et al., J. Pharmaceut. Sci., 93(3):601-611 (2004), E. C.
  • a typical, non-limiting, process of preparing a solvate would involve dissolving a compound of the invention in a desired solvent, which may be water, an organic solvent or a mixture thereof at temperatures above about 20.degree. C, e.g. at room temperature or heating to a temperature above room temperature if appropriate to dissolve the solid, followed by cooling the solution at a rate sufficient to form crystals, and isolating crystals by known methods.
  • a desired solvent which may be water, an organic solvent or a mixture thereof at temperatures above about 20.degree. C, e.g. at room temperature or heating to a temperature above room temperature if appropriate to dissolve the solid, followed by cooling the solution at a rate sufficient to form crystals, and isolating crystals by known methods.
  • a compound of the invention can be recrystallized from an appropriate solvent (e.g., water) to obtain a solvate (e.g., a hydrate).
  • an appropriate solvent e.g., water
  • a solvate e.g., a hydrate
  • Compounds of the invention may contain chemical groups (acidic or basic groups) that can be in the form of salts.
  • Exemplary acid addition salts include acetates (such as those formed with acetic acid or trihaloacetic acid, for example, trifluoroacetic acid), adipates, alginates, ascorbates, aspartates, benzoates, benzenesulfonates, bisulfates, borates, butyrates, citrates, camphorates, camphorsulfonates, cyclopentanepropionates, digluconates, dodecylsulfates, ethanesulfonates, fumarates, glucoheptanoates, glycerophosphates, hemisulfates, heptanoates, hexanoates, hydrochlorides (formed with hydrochloric acid), hydrobromides (formed with hydrogen bromide), hydroiodides, 2-hydroxyethanesulfonates, lac
  • Exemplary basic salts include ammonium salts, alkali metal salts such as sodium, lithium, and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases (for example, organic amines) such as benzathines, dicyclohexylamines, hydrabamines [formed with N,N-bis(dehydro-abietyl)ethylenediamine], N-methyl-D- glucamines, N-methyl-D-glucamides, t-butyl amines, and salts with amino acids such as arginine, lysine and the like.
  • organic bases for example, organic amines
  • organic bases for example, organic amines
  • benzathines dicyclohexylamines
  • hydrabamines [formed with N,N-bis(dehydro-abietyl)ethylenediamine]
  • N-methyl-D- glucamines N-methyl-D-glucamides
  • Basic nitrogen-containing groups may be quaternized with agents such as lower alkyl halides (e.g., methyl, ethyl, propyl, and butyl chlorides, bromides and iodides), dialkyl sulfates (e.g., dimethyl, diethyl, dibutyl, and diamyl sulfates), long chain halides (e.g., decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides), aralkyl halides (e.g., benzyl and phenethyl bromides), and others.
  • lower alkyl halides e.g., methyl, ethyl, propyl, and butyl chlorides, bromides and iodides
  • dialkyl sulfates e.g., dimethyl, diethyl, dibutyl, and diamyl sulfates
  • Salts of the invention include “pharmaceutically acceptable salts” which refers to those salts which retain the biological effectiveness and properties of the free bases or free acids, and which are not biologically or otherwise undesirable.
  • Pharmaceutically acceptable salts comprise pharmaceutically-acceptable anions and/or cations.
  • inventive compounds may have trans and cis isomers and may contain one or more chiral centers, therefore exist in enantiomeric and diastereomeric forms.
  • the invention includes all such isomers, as well as mixtures of cis and trans isomers, mixtures of diastereomers and racemic mixtures of enantiomers (optical isomers).
  • optical isomers When no specific mention is made of the configuration (cis, trans or R or S) of a compound (or of an asymmetric carbon), then any one of the isomers or a mixture of more than one isomer is intended.
  • the processes for preparation can use racemates, enantiomers, or diastereomers as starting materials.
  • enantiomeric or diastereomeric products When enantiomeric or diastereomeric products are prepared, they can be separated by conventional methods, for example, by chromatographic or fractional crystallization.
  • the inventive compounds may be in the free or hydrate form.
  • the atoms therein may have various isotopic forms, e.g., isotopes of hydrogen include deuterium and tritium. All isotopic variants of compounds of the invention are included within the invention and particularly included at deuterium and 13C isotopic variants. It will be appreciated that such isotopic variants may be useful for carrying out various chemical and biological analyses, investigations of reaction mechanisms and the like. Methods for making isotopic variants are known in the art.
  • the bipyDCs represent a class of compounds for the development of antifibrotic or antimetastatic therapeutics.
  • these compounds possess a variety of undesirable chemical properties that have limited their development thus far.
  • the bipyDCs as a class are not cell permeable, requiring the preparation of suitable cell permeable prodrugs.
  • the bipyDCs as a class are not cell permeable, requiring the preparation of suitable cell permeable prodrugs.
  • bipyDCs are capable of binding and forming complexes with free iron and likely other biologically relevant metals.
  • the present work initially focused on possible improvements of
  • Vasta etal. 2016 and the supplemental information for this paper provide additional experimental details including HPLC data, NMR and complex absorbtion spectra, Job's plots and pH titration curves. This reference and the supplemental information are each incorporated by reference herein in its entirety for such details.
  • bipy scaffold itself was problematic and proceeded to assess alternative biheteroaryl scaffolds for potent and selective inhibition of human CP4H, but with improved solubility and iron binding properties.
  • one of pyridine rings of bipy was replaced with another 5- or 6-membered aromatic ring containing an alkaloid nitrogen.
  • replacement of one pyridine with various 5- membered ring heterocycles was assessed.
  • a library of biheteroaryl compounds ( Figure 2) was prepared and evaluated them as iron chelators in vitro.
  • the library was assembled either from commercial vendors (pypyr and pyim) or by synthesis using palladium-catalyzed cross-coupling reactions (pypyraz, pypyrid, pyox, and pythi).
  • pypyraz, pypyrid, pyox, and pythi palladium-catalyzed cross-coupling reactions
  • the Fe2o-EC5o values of the biheteroaryl compounds varied by more than 2 orders of magnitude ( Figures 3A and 3B; Table 2). All, however, were significantly greater than that of bipy or bipyDCs. 38 The Fe2o-EC5o value appeared to rely on both the ring size and pK a of the conjugate acid of the alternative heterocycle, as scaffolds with five-membered rings were substantially weaker chelators than were those with six membered rings, and the pK a value correlated positively with iron affinity.
  • pyox 0.8 (54) ⁇ 3 NO g NO g >18,000 a pK a value of the conjugate acid of pyridine (bipy) or the nonpyridyl ring.
  • b pK a value of the conjugate acid as estimated by titration of the entire compound; bipy, ref 55. determined using spectrophotometry (see The Examples). d Determined by Job's method. 56 ' 57
  • biheteroaryl dicarboxylates were then assessed as inhibitors of human CP4H1.
  • previously described assay conditions (10 ⁇ compound and 50 ⁇ FeS04) in which potent chelators like bipy do not cause inhibition 38 were used.
  • Figure 4 In an initial screen ( Figure 4), some biheteroaryl dicarboxylates showed little or no inhibition of human CP4H1, consistent with the inability of their heteroatoms to participate in an enzymic interaction.
  • imidazole exists as two tautomers, one with a proton on Nl (as in the depiction of pyimDC in Figure 2) and another with a proton on N3.
  • complex formation was not observe between pyimDC and free iron by spectrophotometry, it was found that pyimDC was able to deter the formation of the Fe(bipy)3 2+ complex in a dose-dependent manner ( Figure 5A and the Table in Figure 5B).
  • competition required a free carboxylate on the imidazole ring.
  • NMe-pyimDC ( Figure 2), that is methylated on Nl.
  • NMe- pyimDC was also able to deter the formation of the Fe(bipy)3 2+ complex, but only at high concentrations (Table in Figure 5B).
  • NMe-pyimDC was found to be an inhibitor of human CP4H in vitro, but that its potency is less than that of pyimDC ( Figure 4).
  • the inhibition curves for pyimDC, pythiDC, and pyoxDC were found to be sigmoidal, yielding IC 50 values in the low- micro molar range ( Figure 6A).
  • bipy45'DC and bipy55'DC 21 ' 38 pyimDC, pyoxDC, and pythiDC could bind in the AKG binding pocket and use their second carboxyl group to form additional interactions with residues in the active site of CP4H. If so, then the biheteroaryl dicarboxylates should exhibit structure- activity relationships similar to those of bipy45'DC and bipy55'DC for the inhibition of PHD2, another P4H enzyme that has been characterized extensively. PyimDC, pythiDC, and pyoxDC inhibit human PHD2 only weakly ( Figure 7), with pythiDC displaying especially low potency.
  • HEK human embryonic kidney.
  • IRPs iron regulatory proteins 1 and 2
  • HIF- ⁇ The stability of HIF- ⁇ is dependent on the prolyl 4-hydroxylase activity of PHD2, which is an activity that is inherently sensitive to the iron status of the cell.
  • PHD2 prolyl 4-hydroxylase activity of PHD2
  • iron-deficient cells exhibit ferritin levels that are lower and TFR and HIF- ⁇ levels that are higher than those of untreated cells.
  • MDA-MB-231 cells were treated with biheteroaryl dicarboxylates and assayed for cytotoxicity and indicators of iron deficiency. None of the esterified biheteroaryl dicarboxylates exhibited cytotoxic activity at high micromolar concentrations (The
  • 2,4-Pyridinedicarboxylic acid (24PDC), 2,5-pyridinedicarboxylic acid (25PDC), 2,2 ' - bipyridine (bipy), 2-(lH-Imidazol-2-yl)pyridine (pyim), and 2,2 ' -bipyridine-5,5 ' - dicarboxylic acid (bipy55 T ) C) were obtained from Sigma- Aldrich (St. Louis, MO).
  • 2,2 ' - bipyridine-4,4 ' -dicarboxylic acid (bipy44T ) C) was obtained from TCI America (Portland, OR).
  • 2-(lH-pyrazol-3-yl)pyridine was from Combi-Blocks (San Diego, CA).
  • Phosphine ligands and phosphonium salts were obtained from either Sigma-Aldrich or Strem
  • Pd(OAc)2 was obtained from Sigma-Aldrich, stored in a dessicator, and used without further purification.
  • Deferroxamine mesylate was obtained from Santa Cruz Biotechnology (Dallas, TX).
  • Ethyl dihydroxybenzoate (EDHB) was obtained from Combi-Blocks (San Diego, CA) and recrystallized from EtOAc before use in cell culture experiments. All other reagent chemicals were obtained from commercial sources (Sigma-Aldrich, Acros, Combi-Blocks, Oakwood Products, Enamine, Bachem, or Novabiochem) and used without further purification.
  • the HIF- 1 a peptide55 6 -575 was from AnaSpec (Fremont, CA) and used without further purification. All glassware was flame- or oven-dried, and reactions were performed under N 2 (g) unless indicated otherwise. DCM and toluene were dried over a column of alumina. Dimethylformamide was dried over alumina and further purified through an isocyanate scrubbing column. Other anhydrous solvents were obtained in septum-sealed bottles. Flash chromatography was performed with columns of 40-63 A silica gel, 230 ⁇ 400 mesh (Silicycle, Quebec City, Canada).
  • Thin-layer chromatography was performed on plates of EMD 250 ⁇ silica 6O-F254 with visualization by UV light or staining with KMn0 4 .
  • concentration under reduced pressure refers to the removal of solvents and other volatile materials using a rotary evaporator at water aspirator pressure ( ⁇ 20 torr) while maintaining water-bath temperature below 40°C. Residual solvent was removed from samples at high vacuum ( ⁇ 0.1 torr).
  • high vacuum refers to vacuum achieved by a mechanical belt-drive oil pump. All reported yields are unoptimized. All procedures were performed at ambient temperature unless indicated otherwise.
  • NMR spectra were acquired at ambient temperature with a Bruker DMX-400 Avance spectrometer or a Avance 500i spectrometer from Brucker (Billerica, MA) at the National Magnetic Resonance Facility at Madison (NMRFAM) and were referenced to TMS or a residual protic solvent.
  • Some compounds exist as either mixtures of rotomers or tautomers that do not interconvert on the NMR timescale at ambient temperature and therefore exhibit multiple sets of NMR signals.
  • Electrospray ionization (ESI) and electron ionization (EI) mass spectrometry were performed with a Micromass LCT ® or Micromass AutoSpec ® instruments, respectively, from Waters (Milford, MA) at the Mass Spectrometry Facility in the
  • Human CP4H containing the oc(I) isoform was produced heterologously in Origami B(DE3) Escherichia coli cells and purified as described previously. 35
  • the catalytic activity of human CP4H1 was assayed as described previously. 35 Briefly, activity assays were carried out at 30 °C in 100 ⁇ L Tris-HCl buffer, pH 7.8, containing human CP4H1 (100 nM), inhibitor (0-500 ⁇ ), substrate (dansylGlyProProGlyOEt, 500 ⁇ ), FeS0 4 (50 ⁇ ), BSA (1 mg/mL), catalase (0.1 mg/mL), sodium ascorbate (2 mM), DTT (100 ⁇ ), and a-ketoglutarate (100 ⁇ ). Reactions were pre-incubated with or without inhibitor for 2 min at 30 °C, after which the reaction was initiated by the addition of oc- ketoglutarate.
  • reactions mixtures were pre-incubated with or without inhibitor for 2 min at 30 °C, after which the reaction was initiated by the addition of oc- ketoglutarate. After 10 minutes, reactions were quenched by boiling for 60 s and centrifuged at 10,000g. The supernatant (50 ⁇ ) was injected into a Nucleodur ® C18 Gravity reversed- phase column (4.6 x 250 mm, 5 ⁇ particle size) from Macherey-Nagel (Bethlehem, PA). The column was eluted at 1 mL/min with a gradient (34 min) of 5-56% aqueous acetonitrile containing 0.1 % v/v TFA.
  • the pKa value of biheteroaryl compounds was estimated with potentiometric titration.
  • the affinity of biheteroaryl ligands for Fe(II) was determined comparatively by measuring the half-maximal concentration (EC 50 ) required for binding 20 ⁇ Fe(II) (Fe 2 o-EC 5 o) in sodium phosphate buffer at pH 7.
  • Stock solutions of ligands were prepared in either water for high affinity ligands (typically Fe 2 o-EC5o ⁇ ImM) or DMSO for low affinity ligands (typically Fe2o-ECso > 1000 ⁇ ).
  • Stock solutions of FeS0 4 were prepared in 3 ⁇ 40 and used within 3 hours of preparation.
  • Ligand solutions (3-18000 ⁇ depending on affinity) were prepared in 10 mM sodium phosphate buffer, pH 7, after which Fe(II) stock solution was added to initiate complex formation.
  • solutions were allowed to equilibrate for 15 min, after which the absorbance was recorded at the for the complex under study.
  • the corresponding Fe(II) complexes were unstable and observed to dissociate over time. Therefore, the absorbance value was determined within 30 s of mixing, where the absorbance was measured at the ⁇ for the complex under study. Complexes with pyox were not observed under these conditions. Absorbance values were corrected by subtracting the absorbance value in the absence of ligand.
  • Dose-response curves were generated for each ligand by plotting the absorbance versus the log of the ligand concentration. Fe 2 o-ECso values for each ligand were interpolated from the dose-response curves by no n- linear regression using the sigmoidal dose-response function in Prism. All experiments were performed in triplicate. Experiments to study pyimDC were performed as described above, except that phosphate buffer was excluded from the assay solution, bipy was added last to a final concentration of 300 ⁇ , and the absorbance of the Fe(bipy) 3 2+ complex was measured at 523 nm.
  • the stoichiometry of biheteroaryl complexes with Fe(II) was estimated via Job's method. Briefly, reactions were prepared such that the total moles of Fe(II) and ligand was kept constant, but the mole fraction of the ligand was varied from 0 to 1. The total concentration of ligand and Fe(II) used for each individual ligand was based upon the iron affinity and extinction coefficient, and ranged from 0.4 mM to 2 mM. Stock solutions of ligands were prepared in water. Stock solutions of FeS0 4 were prepared in water and used within 3 hours of preparation. Reactions were prepared in 10 mM sodium phosphate buffer, pH 7.0, and complex formation was initiated by the addition of Fe(II) solution.
  • Job's plots were constructed by plotting the normalized absorbance versus the mole fraction of biheteroaryl ligand, after which the stoichiometry of the complex was estimated from the mole fraction of the reaction with the highest absorbance value. If necessary, blank titrations using only Fe(II) were used to correct the Job's plots.
  • the HEK293T cell line was obtained from American Type Culture Collection (ATCC, Manassas, VA), and the MDA-MB-231 cell line was a generous gift from Dr. Beth Weaver. Cell lines were maintained according to the procedures recommended by the ATCC. Cells were grown in a cell culture incubator at 37 °C under CO2 (5% v/v) in flat-bottomed culture flasks. The culture medium was DMEM supplemented with GIBCO fetal bovine serum (FBS) (10% v/v), penicillin (100 units/mL), streptomycin (100 ⁇ g/mL) and L-glutamine (2 mM). Cells were counted by hemocytometry with Trypan Blue prior to use in assays. XI. Cytotoxicity Assays
  • MDA-MB-231 cells grown as described in Section X were plated at a concentration of 5,000 cells/well in a clear 96-well plate. The cells were allowed to adhere for 4 h, after which the medium was removed and discarded. Fresh medium was added and the cells were treated with varying concentrations of the test compound at 37 °C for 24 h. The medium was removed, and cells were washed with Dulbecco's PBS.
  • the MTS reagent was added at a ratio of 1:5, and the cells were incubated at 37 °C for 2 h before measuring the absorbance at 490 nm. The average absorbance was measured in triplicate for each concentration tested, and the entire experiment was repeated in duplicate. The percentage of viable cells was determined by normalizing to a PBS control (100% viable), and a H2O2 control (0% viable). For all of the compounds tested, the LD5 0 - value could not be measured due to its being higher than the aqueous solubility limit: diethyl bipy55'DC, LD5 0 >100 ⁇ ; diethyl pyimDC, LD 50 >1000 ⁇ ; diethyl pythiDC, LD 50 >500 ⁇ .
  • XII Effect of CP4H Inhibitors on Iron Metabolism and P4HA1 Levels in Human Cells
  • MDA-MB-231 or HEK293T cells were plated at 50,000 cells/well into 12-well plates and grown to confluence (-24 h) as described in Section X. The cells were washed, and the medium was replaced with 1.0 mL of serum- free medium. Stock solutions of all test compounds were prepared at lOOx in DMSO and added to a final concentration of lx (0.1- 0.5 mM). After addition of the test compound or DMSO vehicle, cells were incubated for 24 h. The cells were then washed with 100 MPER solution from Thermo Fisher Scientific (Waltham, MA) and collected. Protein concentrations were determined by BCA assay (Thermo Fisher Scientific), and samples corresponding to 40 ⁇ g total protein were analyzed by immunoblot.
  • XIII Immuno blotting
  • the anti-rabbit antibody from Promega and the anti-mouse antibody from Abbiotec (San Diego, CA) were used at the working dilution specified by the manufacturer.
  • Statistical comparisons were performed using Student's i-test or one-way ANOVA functions available in Prism.
  • P4HA1 levels protein samples were treated and quantified as described above for iron metabolism except that the blots were probed with primary antibodies to human P4HA1 (rabbit polyclonal) from Thermo Fisher Scientific and human ⁇ -actin.
  • MDA-MB-231 cells were plated at 3 x 10 6 cells/dish into 100 mm x 20 mm dishes and grown to confluence (-24 h) as described in Section X. The cells were washed, and the medium was replaced with 10 mL of serum-free medium. Stock solutions of all test compounds were prepared at lOOx in DMSO and added to a final concentration of lx (as denoted). After addition of the test compound or DMSO vehicle, cells were incubated for 24 h, after which they were harvested by trypsinization and centrifugation. The resultant pellets (-50 ⁇ ) were washed with PBS and then lysed using a protocol described previously.
  • cell pellets were resuspended in 200 of cell lysis buffer, which was 20 mM Hepes-HCl buffer, pH 7.4, containing sodium pyrophosphate (10 mM), sodium fluoride (50 mM), ⁇ -glycerophosphate (50 mM), EDTA (5 mM), GTP (1 mM), sodium orthovanadate (1 mM), benzamidine hydrochloride (2 mM), Nonidet NP-40 (0.5% v/v), p-nitrophenyl p'- guanidinobenzoate-HCl (25 ⁇ g/mL), DTT (1 mM), leupeptin (40 ⁇ g/mL), pepstatin (4 ⁇ g/mL), SBTI (100 ⁇ g/mL), MG132 (10 ⁇ ), PMSF (200 ⁇ ), and BHT (5 ⁇ g/mL) by vortexing and lysed on ice by vortexing every few min. After 15 min, the lys
  • MDA-MB-231 cells were plated at 50,000 cells/well into 12-well plates and grown to confluence (-24 h) as described in Section X. The cells were washed, and the medium was replaced with 1.5 mL of serum- free medium containing sodium ascorbate (50 ⁇ g/mL). Stock solutions of all test compounds were prepared at lOOx in DMSO and added to a final concentration of lx (as denoted). After the addition of a test compound or vehicle (DMSO), cells were incubated for 48 h, after which 1.0 mL of conditioned medium was removed and added to 4.0 mL of chilled acetone (-20 °C).
  • DMSO test compound or vehicle
  • the resultant mixtures were incubated for 3 h at -20 °C, after which the precipitated protein was pelleted by centrifugation at 10,000g for 30 min. The supernatants were discarded and the pellets dried briefly in a fume hood. Pellets were stored at -20 °C prior to analysis with an immunoblot.
  • trimethylsilylacetylene (420 ⁇ , 3.0 mmoles) in DCM (5 mL) was added to the flask while stirring on ice.
  • Triethylamine (1.4 mL, 10.0 mmoles) was added the flask while stirring, after which the flask was allowed to come to room temperature overnight.
  • the reaction mixture was diluted with hexanes (5 mL) and filtered through Celite ® , and the filtrate was washed with H20 (50 mL) and brine (50 mL).
  • reaction mixture was cooled and filtered through Celite ® , and the filtrate was concentrated under reduced pressure.
  • the crude product was then further purified by chromatography on silica (45 % EtOAc in hexanes) to afford the title compound (100 mg) as a pale orange solid. Due to the presence of minor contaminants that were difficult to remove by chromatography or recrystallization, the slightly crude product was used directly in the next reaction before further purification and characterization.
  • Ethyl 2-(5-ethoxycarbonyl-lH-pyrrol-3-yl)pyridine-5-carboxylate Procedure: Ethyl 2-ethynylpyridine-5-carboxylate (91 mg, 0.52 mmoles) was dissolved in toluene (5 mL) in a glass vial and the solution was chilled on ice with stirring. A 73 % (w/w) solution of ethyl diazoacetate in DCM (162 mg, 1.04 mmoles) was added dropwise, after which the vial was purged with N 2 (g) and stirred at 95 °C.
  • the aqueous layer was extracted with DCM (4 x 30 mL) and the combined organics were dried over Na 2 S0 4 (s) and concentrated under reduced pressure to afford the crude alcohol (678 mg) as a pale yellow solid.
  • the crude alcohol was dissolved in dry DCM (45 mL), after which Dess-Martin periodinane (2.6 g, 6.1 mmoles) was added portion wise while stirring on ice. After 6 h, the reaction was quenched by the dropwise addition of a solution of 5% Na 2 S 2 (3 ⁇ 4 in half saturated NaHC0 3 (80 mL). The aqueous layer was extracted with DCM (3 x 40 mL).
  • HGlyOEt- HC1 (333 mg, 2.4 mmoles) and DMAP (29.1 mg, 0.24 mmoles) were added to a dried flask. The flask was capped with a septum and purged with nitrogen ( ⁇ 5 times). DCM (6 mL) was added and the reaction was cooled in an ice bath. DIEA (750 ⁇ , 4.3 mmoles) and monomethyl oxalyl chloride (200 ⁇ , 2.2 mmoles) were added via syringe, and the reaction was stirred and allowed to come to room temperature. After 6 hr, the reaction was quenched on ice by the dropwise addition of saturated ammonium chloride (10 mL).
  • N-(Methoxyoxalyl)glycine ethyl ester 100 mg, 0.53 mmoles
  • KOH 88 mg, 1.6 mmoles
  • MeOH 5.0 mL
  • the reaction mixture was heated to 60 °C with stirring until the starting material was completely consumed, as judged by TLC.
  • the reaction mixture was cooled and concentrated under reduced pressure.
  • the crude product was dissolved in water (2 mL), and the pH adjusted to

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  • Organic Chemistry (AREA)
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Abstract

La présente invention concerne des esters et des dicarboxylates bihétéroaryliques, et des sels de ceux-ci qui sont utiles comme modulateurs de l'activité de la CP4H et plus particulièrement comme inhibiteurs de la CP4H. La présente invention concerne les composés de formule (I), et des sels de ceux-ci, dans laquelle : X représente S, O, NH, ou NR, R représentant un groupe alkyle possédant de 1 à 3 atomes de carbone ; R1 et R2 représentent indépendamment -OR7, ou -NHSO2R8, R7 étant choisi parmi : un hydrogène, un alkyle, un alcényle, un alcoxyalkyle, -R'-CO-R", -R'-CO-O-R", -CO-R", -R'-O-CO-R", -R'-CO-NR", -CO-NR", ou -R'-O-CO-NR", et R8 étant choisi parmi un hydrogène, un alkyle, un aryle, un arylalkyle ; R3, R4 et R6 représentent indépendamment un hydrogène, un alkyle, un alcoxy, un alcényle, un alcénoxy, un haloalkyle, un haloalcényle, un halogène, un hydroxyle, un hydroxyalkyle, un hydroxyalcényle, un aryle, un aryloxy, un arylalkyle ou un arylalkyloxy ; R5 représente un hydrogène, un halogène, un alkyle possédant de 1 à 3 atomes de carbone, ou un alcoxy possédant de 1 à 3 atomes de carbone ; -R'- représente un alkylène ramifié ou à chaine droite divalente, et -R" représente un groupe alkyle, alcényle, arylalkyle, ou aryle. L'invention concerne également des méthodes d'inhibition de la CP4H in vivo et in vitro.
PCT/US2016/024522 2015-03-28 2016-03-28 Inhibiteurs de la collagène prolyl-4-hydroxylase WO2016160706A1 (fr)

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CA3088150A1 (fr) * 2018-01-09 2019-07-18 Cornell University Prevention et traitement de fibrose d'organe
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Citations (3)

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US4904675A (en) * 1987-02-10 1990-02-27 Zyma Sa Phamacologically active 5-carboxy-2-(5-tetrazolyl) pyridines
US5614520A (en) * 1990-11-30 1997-03-25 Teijin Limited 2-arylthiazole derivatives and pharmaceutical composition thereof
US20130065918A1 (en) * 2011-03-10 2013-03-14 Boehringer Ingelheim International Gmbh Soluble guanylate cyclase activators

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US4904675A (en) * 1987-02-10 1990-02-27 Zyma Sa Phamacologically active 5-carboxy-2-(5-tetrazolyl) pyridines
US5614520A (en) * 1990-11-30 1997-03-25 Teijin Limited 2-arylthiazole derivatives and pharmaceutical composition thereof
US20130065918A1 (en) * 2011-03-10 2013-03-14 Boehringer Ingelheim International Gmbh Soluble guanylate cyclase activators

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Title
GILKES ET AL.: "Collagen Prolyl Hydroxylases Are Essential for Breast Cancer Metastasis", MOLECULAR AND CELLULAR PATHOBIOLOGY, CANCER RESEARCH, vol. 73, no. 11, 2013, pages 3285 - 3296, XP055317453 *
HALES ET AL.: "Novel Inhibitors of prolyl 4-hydroxylase. 5. The intriguing structure-activity relationships seen with 2,2'-bipyridine and its 5,5'-dicarboxylic acid derivatives", J. MED. CHEM., vol. 36, no. 24, 1993, pages 3853 - 3858, XP002313833 *

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